Throughout this application, various patents, published patent applications and publications, are referenced. Disclosures of these patents, published patent applications and publications, in their entireties, are hereby incorporated by reference into this application. Included among the patents and applications incorporated by reference are U.S. Provisional Patent Application 60/782,515 and copending International Application No. PCT/SG2007/000071 (which designates the United States). In the case of conflict, the present specification, including definitions, will control. Full bibliographic citations for the publications may be found listed in the List of References immediately preceding the claims.
Natural products (NPs) are typical secondary metabolites produced by organisms in response to external stimuli, such as nutritional changes, infection, and adaptive evolution. Several different NPs produced by plants, fungi, bacteria, protozoans, insects and animals have been isolated as biologically active pharmacophores. Well-known examples of valuable NPs used widely in medical and animal health industry include lovastatin (anticholesterolemic agent), cyclosporine A and tacrolimus (immunosuppressive agents), paclitaxel and doxorubicin (antitumor agents), erythromycin (antibiotic), and amphotericin B (fungicidal agent) (Strohl 2000).
A wide variety of actinomycetes have been shown to exhibit significant antifungal activity (Lee and Hwang 2002). Likewise, filamentous fungi are also known to produce a variety of antifungal compounds, including echinocandins, ergokinin A, sphingofungin, peptaibols, and several other compounds with a diversity of core structures. A variety of pseudomonads have been shown to synthesize seed- and crop-protecting antifungals like pyrrolnitrin, syringomycin etc (Rangaswamy et al, 1998). Similarly, extracts of many plants have been shown to contain low-molecular-weight compounds, which exhibit antifungal activity in vitro. These compounds include a diverse array of secondary metabolites, such as phenolics, saponins, cyanogenic glycosides, cyclic hydroxamic acids, sesquiterpenes, isoflavonoids, sulfur-containing indole derivatives, and many other compounds (Osbourn, 1999). Flocculosin is a novel low-molecular-weight glycolipid isolated from the yeast-like fungus Pseudozyma flocculosa. It is used to control fungal powdery mildew disease in plants and has also been successfully tested against human fungal pathogens like C. albicans and Cryptococcus neoformans (Mimee et al, 2005).
In spite of the progress in antifungal therapy, drugs like amphotericin B or triazole have limited use because of their toxicity and/or drug resistance issues (Bagnis and Deray, 2002). Other promising candidate drugs like Caspofungin have low oral bioavailability (Boucher et al, 2004). Hence, there is a need for the isolation or synthesis of new compounds with different modes of action and low toxicity.
ATP-binding cassette (ABC) transporters, which constitute the largest superfamily of proteins known, are able to couple the hydrolysis of ATP to the transport of a variety of substrates either into or out of cells (Ritz et al. 2003). In humans, loss of ABC transporter function has been implicated in several pathologies including cystic fibrosis, cholestasis, artherosclerosis, hypoglycemia, hyperbiliruginemia, and macular dystrophy and degenerative diseases (Pastan and Gottesmann 1988). Moreover, the P-glycoprotein class of ABC transporters is able to efflux chemotherapeutic drugs and lipids, resulting in reduced effectiveness of cancer treatments (Tsuruo et al, 2003). Similarly, ABC transporters in bacteria are essential for survival and are also required to secrete toxins and antimicrobial agents (Buchaklian and Klug, 2006).
Loss-of-function analysis of ABC3, which encodes a novel multidrug resistance transporter in the cereal pathogen Magnaporthe grisea, showed that MDR-based efflux plays an essential role in fungal pathogenesis (Sun et al. 2006; PCT International Patent Application No. PCT/SG2007/000071). Abc3-deletion strain of M. grisea has been classified as a non-pathogenic mutant. Although it forms the infection structures called appressoria, the lack of infectivity in the abc3-delete mutant was correlated to its inability to penetrate the host tissue, which in turn, was proposed to be due to accumulation of an inhibitory metabolite and/or perturbed redox homeostasis within the appressoria. Further characterization confirmed that Abc3 function is required by the blast fungus to withstand cytotoxic and oxidative stress especially within the appressoria during infection.